the objective of this project is to use affinity labeling as a tool for comparing the active sites of glycolytic and gluconeogenic enzymes which use either 2-phosphoglyceric acid or phosphoenolpyruvate. These enzymes (phosphoglycerate mutase, diphosphoglycerate mutase, enolase, pyruvate kinase, PEP carboxylase, and PEP carboxykinase) may be amenable to affinity labeling by reagents which contain a carboxyl and a phosphate group as affinity groups. Toward this goal, two potential affinity labels for the above enzymes will be synthesized, one containing a reactive azide group, the other a bromoacetyl group. If either of these compounds is a successful affinity label, the site(s) of reaction on the enzyme(s) will be explored. These two reagents will also be tested as ligands for affinity chromatography systems for the above enzymes. Another reagent which will be used is glycidol phosphate (1,2 epoxypropanol 3-phosphate), which has been demonstrated to be an affinity label for both enolase and triose phosphate isomerase. The site of reaction of glycidol phosphate with rabbit muscle enolase will be determined, for comparison with the active site peptide of triose phosphate isomerase (determined by other laboratories).